Reduction of erythrocyte sedimentation rate

ABSTRACT

The present invention relates to reduction of erythrocyte sedimentation rate in a blood sample. In particular, formulations, compositions, articles of manufacture, kits and methods for reduced erythrocyte sedimentation rate in a blood sample are provided.

CROSS-REFERENCE

This application is a continuation of U.S. patent application Ser. No.15/750,590, filed May 31, 2018, now allowed, which is a § 371 U.S.National Entry of PCT/US2016/065198, filed Dec. 6, 2016, which claimsthe benefit of US Provisional Application No. 62/264,786, filed Dec. 8,2015, each of which is incorporated by reference herein in its entirety.

BACKGROUND OF THE INVENTION

There exists a need for improved formulations and methods for reducingthe rate of blood sedimentation for a time sufficient for storage,transport, and shipping for research, diagnostic and therapeuticpurposes.

The present invention relates generally to the reduction ofsedimentation rate of one or more erythrocytes in a blood sample. Inparticular, the invention relates to formulations, compositions,articles of manufacture, kits and methods for the reduction oferythrocyte sedimentation rate in a blood sample.

SUMMARY OF THE INVENTION

Described herein, in some embodiments, are in vitro methods for reducingthe erythrocyte sedimentation rate in a blood sample, comprising:combining a sample of blood with an amount of a formulation comprisingsucralose, wherein the amount is sufficient to produce a treated bloodsample having a sucralose concentration of at least about 5 mMsucralose, thereby reducing the erythrocyte sedimentation rate ascompared to erythrocyte sedimentation rate in an untreated blood sample.In some embodiments, the treated blood sample has a sucraloseconcentration of about 5 mM sucralose to about 100 mM sucralose. In someembodiments, the treated blood sample has a sucralose concentration ofabout 10 mM sucralose to about 100 mM sucralose. In some embodiments,the treated blood sample has a sucralose concentration of about 20 mMsucralose to about 100 mM sucralose. In some embodiments, the treatedblood sample has a sucralose concentration of about 24 mM sucralose toabout 100 mM sucralose. In some embodiments, the treated blood samplehas a sucralose concentration of about 15 mM sucralose to about 50 mMsucralose. In some embodiments, the treated blood sample has a sucraloseconcentration of about 5 mM sucralose to about 50 mM sucralose. In someembodiments, the treated blood sample has a sucralose concentration ofabout 5 mM sucralose to about 40 mM sucralose. In some embodiments, thetreated blood sample has a sucralose concentration of about 10 mMsucralose to about 35 mM sucralose. In some embodiments, the treatedblood sample has a sucralose concentration of about 10 mM sucralose toabout 30 mM sucralose. In some embodiments, the treated blood sample hasa sucralose concentration of about 5 mM sucralose to about 25 mMsucralose. In some embodiments, the treated blood sample has a sucraloseconcentration of about 5 mM sucralose up to but not including 25 mMsucralose. In some embodiments, the treated blood sample has a sucraloseconcentration of about 5 mM sucralose to about 20 mM sucralose. In someembodiments, the treated blood sample has a sucralose concentration ofabout 5 mM sucralose to about 15 mM sucralose. In some embodiments, thetreated blood sample has a sucralose concentration of about 5 mMsucralose to about 10 mM sucralose. In some embodiments, the treatedblood sample has a sucralose concentration of about 24 mM. In someembodiments, the treated blood sample has a sucralose concentration ofabout 25 mM. In some embodiments, erythrocyte sedimentation rate isreduced by at least 10%, at least 20%, at least 30%, at least 40%, atleast 50%, at least 60%, at least 70%, at least 80%, at least 90%, atleast 95% of the erythrocyte sedimentation rate of the untreated bloodsample. In some embodiments, the formulation is in the form of a powder,a solid, a lyophilized form, a solution, or an aqueous solution. In someembodiments, the formulation is a powder. In some embodiments, theformulation is a solid. In some embodiments, the formulation islyophilized. In some embodiments, the formulation is a solution. In someembodiments, the solution is an aqueous solution. In some embodiments,the formulation consists of sucralose. In some embodiments, theformulation further comprises an anticoagulant. In some embodiments, theanticoagulant is selected from the group consisting of acid citratedextrose solution (ACD), sodium heparin, sodium fluoride, lithiumheparin, tri-potassium ethylenediaminetetraacetic acid (K₃EDTA),di-potassium ethylenediaminetetraacetic acid (K₂EDTA), hirudin, andsodium polyanethol sulfonate (SPS). In some embodiments, theanticoagulant is acid citrate dextrose solution (ACD). In someembodiments, the anticoagulant is sodium heparin. In some embodiments,the anticoagulant is sodium fluoride. In some embodiments, theanticoagulant is lithium heparin. In some embodiments, the anticoagulantis tri-potassium ethylenediaminetetraacetic acid (K₃EDTA). In someembodiments, the anticoagulant is di-potassiumethylenediaminetetraacetic acid (K₂EDTA). In some embodiments, theanticoagulant is hirudin. In some embodiments, the anticoagulant issodium polyanethol sulfonate (SPS). In some embodiments, the formulationis contained within a blood collection tube, and the combining stepoccurs within the blood collection tube. In some embodiments, the bloodcollection tube is an evacuated blood collection tube. In someembodiments, the blood is collected from a subject. In some embodiments,the subject is a mammal. In some embodiments, the subject is a human.

Described herein, in some embodiments, are in vitro methods formaintaining one or more erythrocytes in suspension in a blood sample,comprising: combining a sample of blood with an amount of a formulationcomprising sucralose, wherein the amount is sufficient to produce atreated blood sample having a sucralose concentration of at least about5 mM sucralose, thereby maintaining the one or more erythrocytes insuspension for a period of at least 30 minutes as compared to anuntreated blood sample. In some embodiments, the one or moreerythrocytes remain in suspension for a period of at least 10 minutes,at least 20 minutes, at least 30 minutes, at least 40 minutes, at least50 minutes, at least 60 minutes, at least 90 minutes, at least 2 hours,at least 3 hours, at least 4 hours, at least 5 hours, at least 6 hours,at least 7 hours, at least 8 hours, at least 10 hours, at least 12hours, at least 24 hours or at least 48 hours. In some embodiments, thetreated blood sample has a sucralose concentration of about 5 mMsucralose to about 100 mM sucralose. In some embodiments, the treatedblood sample has a sucralose concentration of about 10 mM sucralose toabout 100 mM sucralose. In some embodiments, the treated blood samplehas a sucralose concentration of about 20 mM sucralose to about 100 mMsucralose. In some embodiments, the treated blood sample has a sucraloseconcentration of about 24 mM sucralose to about 100 mM sucralose. Insome embodiments, the treated blood sample has a sucralose concentrationof about 15 mM sucralose to about 50 mM sucralose. In some embodiments,the treated blood sample has a sucralose concentration of about 5 mMsucralose to about 50 mM sucralose. In some embodiments, the treatedblood sample has a sucralose concentration of about 5 mM sucralose toabout 40 mM sucralose. In some embodiments, the treated blood sample hasa sucralose concentration of about 10 mM sucralose to about 35 mMsucralose. In some embodiments, the treated blood sample has a sucraloseconcentration of about 10 mM sucralose to about 30 mM sucralose. In someembodiments, the treated blood sample has a sucralose concentration ofabout 5 mM sucralose to about 25 mM sucralose. In some embodiments, thetreated blood sample has a sucralose concentration of about 5 mMsucralose up to but not including 25 mM sucralose. In some embodiments,the treated blood sample has a sucralose concentration of about 5 mMsucralose to about 20 mM sucralose. In some embodiments, the treatedblood sample has a sucralose concentration of about 5 mM sucralose toabout 15 mM sucralose. In some embodiments, the treated blood sample hasa sucralose concentration of about 5 mM sucralose to about 10 mMsucralose. In some embodiments, the treated blood sample has a sucraloseconcentration of about 24 mM. In some embodiments, the treated bloodsample has a sucralose concentration of about 25 mM. In someembodiments, at least 10%, at least 20%, at least 30%, at least 40%, atleast 50%, at least 60%, at least 70%, at least 80%, at least 90%, atleast 95% of the one or more erythrocytes remain in suspension in thetreated blood sample as compared to the untreated blood sample. In someembodiments, the formulation is a powder. In some embodiments, theformulation is a solid. In some embodiments, the formulation islyophilized. In some embodiments, the formulation is a solution. In someembodiments, the solution is an aqueous solution. In some embodiments,the formulation consists of sucralose. In some embodiments, theformulation further comprises an anticoagulant. In some embodiments, theanticoagulant is selected from the group consisting of acid citratedextrose solution (ACD), sodium heparin, sodium fluoride, lithiumheparin, tri-potassium ethylenediaminetetraacetic acid (K₃EDTA),di-potassium ethylenediaminetetraacetic acid (K₂EDTA), hirudin, andsodium polyanethol sulfonate (SPS). In some embodiments, theanticoagulant is acid citrate dextrose solution (ACD). In someembodiments, the anticoagulant is sodium heparin. In some embodiments,the anticoagulant is sodium fluoride. In some embodiments, theanticoagulant is lithium heparin. In some embodiments, the anticoagulantis tri-potassium ethylenediaminetetraacetic acid (K₃EDTA). In someembodiments, the anticoagulant is di-potassiumethylenediaminetetraacetic acid (K₂EDTA). In some embodiments, theanticoagulant is hirudin. In some embodiments, the anticoagulant issodium polyanethol sulfonate (SPS). In some embodiments, the formulationis contained within a blood collection tube, and the combining stepoccurs within the blood collection tube. In some embodiments, the bloodcollection tube is an evacuated blood collection tube. In someembodiments, the blood sample is collected from a subject. In someembodiments, the subject is a mammal. In some embodiments, the subjectis a human.

Described herein, in some embodiments, are compositions comprising ablood sample and sucralose, wherein the sucralose is at a concentrationof about 5 mM sucralose up to about 100 mM sucralose. In someembodiments, the sucralose is at a concentration of about 10 mMsucralose to about 100 mM sucralose. In some embodiments, the sucraloseis at a concentration of about 20 mM sucralose to about 100 mMsucralose. In some embodiments, the sucralose is at a concentration ofabout 24 mM sucralose to about 100 mM sucralose. In some embodiments,the sucralose is at a concentration of about 15 mM sucralose to about 50mM sucralose. In some embodiments, the sucralose is at a concentrationof about 5 mM sucralose to about 50 mM sucralose. In some embodiments,the sucralose is at a concentration of about 5 mM sucralose to about 40mM sucralose. In some embodiments, the treated blood sample has asucralose concentration of about 10 mM sucralose to about 35 mMsucralose. In some embodiments, the treated blood sample has a sucraloseconcentration of about 10 mM sucralose to about 30 mM sucralose. In someembodiments, the sucralose is at a concentration of about 5 mM sucraloseup to but not including 25 mM sucralose. In some embodiments, thesucralose is at a concentration of about 5 mM sucralose to about 20 mMsucralose. In some embodiments, the sucralose is at a concentration ofabout 5 mM sucralose to about 15 mM sucralose. In some embodiments, thesucralose is at a concentration of about 5 mM sucralose to about 10 mMsucralose. In some embodiments, the treated blood sample has a sucraloseconcentration of about 24 mM. In some embodiments, the treated bloodsample has a sucralose concentration of about 25 mM. In someembodiments, the compositions further comprise an anticoagulant. In someembodiments, the anticoagulant is selected from the group consisting ofacid citrate dextrose solution (ACD), sodium heparin, sodium fluoride,lithium heparin, tri-potassium ethylenediaminetetraacetic acid (K₃EDTA),di-potassium ethylenediaminetetraacetic acid (K₂EDTA), hirudin, andsodium polyanethol sulfonate (SPS). In some embodiments, theanticoagulant is acid citrate dextrose solution (ACD). In someembodiments, the anticoagulant is sodium heparin. In some embodiments,the anticoagulant is sodium fluoride. In some embodiments, theanticoagulant is lithium heparin. In some embodiments, the anticoagulantis tri-potassium ethylenediaminetetraacetic acid (K₃EDTA). In someembodiments, the anticoagulant is di-potassiumethylenediaminetetraacetic acid (K₂EDTA). In some embodiments, theanticoagulant is hirudin. In some embodiments, the anticoagulant issodium polyanethol sulfonate (SPS). In some embodiments, the compositionis contained within a blood collection tube. In some embodiments, theblood collection tube is an evacuated blood collection tube. In someembodiments, the blood is collected from a subject. In some embodiments,the subject is a mammal. In some embodiments, the subject is a human.

Described herein, in some embodiments, are articles of manufacture,comprising sucralose contained within a blood collection tube, whereinthe sucralose is in a quantity sufficient to produce a finalconcentration of about 5 mM sucralose to about 100 mM sucralose in theblood sample. In some embodiments, the sucralose is in a quantitysufficient to produce a final concentration of about 10 mM sucralose toabout 100 mM sucralose. In some embodiments, the sucralose is in aquantity sufficient to produce a final concentration of about 20 mMsucralose to about 100 mM sucralose. In some embodiments, the sucraloseis in a quantity sufficient to produce a final concentration of about 24mM sucralose to about 100 mM sucralose. In some embodiments, thesucralose is in a quantity sufficient to produce a final concentrationof about 15 mM sucralose to about 50 mM sucralose. In some embodiments,the sucralose is in a quantity sufficient to produce a finalconcentration of about 5 mM sucralose to about 50 mM sucralose in theblood sample. In some embodiments, the sucralose is in a quantitysufficient to produce a final concentration of about 5 mM sucralose toabout 40 mM sucralose in the blood sample. In some embodiments, thetreated blood sample has a sucralose concentration of about 10 mMsucralose to about 35 mM sucralose. In some embodiments, the treatedblood sample has a sucralose concentration of about 10 mM sucralose toabout 30 mM sucralose. In some embodiments, the sucralose is in aquantity sufficient to produce a final concentration of about 5 mMsucralose to up to but not including 25 mM sucralose in the bloodsample. In some embodiments, the sucralose is in a quantity sufficientto produce a final concentration of about 5 mM sucralose to about 20 mMsucralose in the blood sample. In some embodiments, the sucralose is ina quantity sufficient to produce a final concentration of about 5 mMsucralose to about 15 mM sucralose in the blood sample. In someembodiments, the sucralose is in a quantity sufficient to produce afinal concentration of about 5 mM sucralose to about 10 mM sucralose inthe blood sample. In some embodiments, the treated blood sample has asucralose concentration of about 24 mM. In some embodiments, the treatedblood sample has a sucralose concentration of about 25 mM. In someembodiments, the sucralose is a powder. In some embodiments, thesucralose is a solid. In some embodiments, the sucralose is lyophilized.In some embodiments, the sucralose is in solution. In some embodiments,the sucralose solution is an aqueous solution. In some embodiments, theblood collection tube is an evacuated blood collection tube. In someembodiments, the articles of manufacture further comprise ananticoagulant. In some embodiments, the anticoagulant is selected fromthe group consisting of acid citrate dextrose solution (ACD), sodiumheparin, sodium fluoride, lithium heparin, tri-potassiumethylenediaminetetraacetic acid (K₃EDTA), di-potassiumethylenediaminetetraacetic acid (K₂EDTA), hirudin, and sodiumpolyanethol sulfonate (SPS). In some embodiments, the anticoagulant isacid citrate dextrose solution (ACD). In some embodiments, theanticoagulant is sodium heparin. In some embodiments, the anticoagulantis sodium fluoride. In some embodiments, the anticoagulant is lithiumheparin. In some embodiments, the anticoagulant is tri-potassiumethylenediaminetetraacetic acid (K₃EDTA). In some embodiments, theanticoagulant is di-potassium ethylenediaminetetraacetic acid (K₂EDTA).In some embodiments, the anticoagulant is hirudin. In some embodiments,the anticoagulant is sodium polyanethol sulfonate (SPS).

Described herein, in some embodiments, are kits, comprising an articleof manufacture provided herein, and a package insert.

INCORPORATION BY REFERENCE

All publications, patents, and patent applications mentioned in thisspecification are herein incorporated by reference to the same extent asif each individual publication, patent, or patent application wasspecifically and individually indicated to be incorporated by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

The novel features of the invention are set forth with particularity inthe appended claims. A better understanding of the features andadvantages of the present invention will be obtained by reference to thefollowing detailed description that sets forth illustrative embodiments,in which the principles of the invention are utilized, and theaccompanying drawings of which:

FIGS. 1A-1C illustrate reduction of erythrocyte sedimentation rate inwhole blood following addition of 0.5 M sucralose, PBS, or saline.Storage was for 0 hours (FIG. 1A), 6 hours (FIG. 1B), and 24 hours (FIG.1C). NF=no formulation control.

FIG. 2 illustrates sedimentation rate of erythrocytes in whole bloodfollowing addition of the indicated solution of sucralose, the indicatedsaccharide, PBS, or saline. Storage was for 1 hour (upper panel) and 6hours (center panel). Lack of hemolysis from the reduction ofsedimentation rate is illustrated in the bottom panel by centrifugationafter 7 hours of storage.

FIG. 3 illustrates sedimentation rate of erythrocytes following additionof sucralose in powder form to the indicated final concentration.Storage of samples was for 1 hour (upper panel) and 6 hours (lowerpanel). NF=no formulation control.

FIG. 4 illustrates the effect of different anticoagulants onsedimentation rate of erythrocytes collected from whole blood. Sampleswere collected into blood collection tubes containing the indicatedanticoagulant. Storage of samples following addition of a solution of0.5 M sucralose or PBS was for 2 hours (upper panel) and 8 hours (lowerpanel). NF=no formulation control.

FIG. 5 illustrates the effect of a solution of 0.5 M sucralose onsedimentation rate of erythrocytes in whole blood compared to PBS, theindicated polyols, and the indicated halogenated polyols. Storage ofsamples was for 2 hours. NF=no formulation control.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to formulations, compositions, articles ofmanufacture, kits, and methods for reduction of sedimentation rate ofone or more erythrocytes in a blood sample.

In some embodiments, the formulations, methods, and compositionsprovided herein provide for reduced sedimentation rate and thus storageof the one or more erythrocytes in a blood sample at the injection siteof a microfluidic device. Reduction of sedimentation rate of one or moreerythrocytes in a blood sample allows for the slow injection of one ormore erythrocytes into the microinjection device without the need forprior sample mixing.

Erythrocyte sedimentation rate is used as a parameter for prognosis ofdiseases such as multiple myeloma, temporal arteritis, polymyalgiarheumatica, systemic lupus erythematosus, and rheumatoid arthritis.Thus, in some embodiments, the formulations, methods, and compositionsfor reduction of erythrocyte sedimentation rate in a blood sampleprovided herein may benefit patients with diseases that correlate withan increased rate of erythrocyte sedimentation.

Definitions

As used in this specification and the appended claims, the singularforms “a”, “an”, and “the” include plural references unless the contextclearly dictates otherwise. Thus, for example, references to “themethod” includes one or more methods, and/or steps of the type describedherein which will become apparent to those persons skilled in the artupon reading this disclosure and so forth.

“About” as used herein when referring to a measurable value such as anamount, a temporal duration, and the like, is meant to encompassvariations of ±20%, or ±10%, or ±5%, or even ±1% from the specifiedvalue, as such variations are appropriate for the disclosed compositionsor to perform the disclosed methods.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of skill in theart to which this invention belongs.

The term “ambient temperature” as used herein refers to common indoorroom temperatures. In some embodiments, ambient temperature is 15 to 32°C. In some embodiments, ambient temperature is 20 to 27° C.

As used herein, the terms “reduced sedimentation rate,” “reducingsedimentation rate,” and “reduction of sedimentation rate,” refer to theability of a material to decrease the sedimentation rate of erythrocytesin a blood sample. In some embodiments, erythrocyte sedimentation rateis reduced by at least 10%, at least 20%, at least 30%, at least 40%, atleast 50%, at least 60%, at least 70%, at least 80%, at least 90%, atleast 95% of the erythrocyte sedimentation rate of the untreated bloodsample. In some embodiments, reduction of sedimentation rate refers tothe ability of a material to prevent one or more erythrocytes in a bloodsample from settling out of suspension due to the force of gravity. Insome embodiments, one or more erythrocytes are maintained in suspensionfor at least 30 minutes. In some embodiments, at least 10%, at least20%, at least 30%, at least 40%, at least 50%, at least 60%, at least70%, at least 80%, at least 90%, at least 95% of one or moreerythrocytes remain in suspension in the treated blood sample ascompared to the untreated blood sample. In some embodiments, one or moreerythrocytes remain in suspension for at least 10 minutes, at least 20minutes, at least 30 minutes, at least 40 minutes, at least 50 minutes,at least 60 minutes, at least 90 minutes, at least 2 hours, at least 3hours, at least 4 hours, at least 5 hours, at least 6 hours, at least 7hours, at least 8 hours, at least 10 hours, at least 12 hours, at least24 hours or at least 48 hours.

Formulation Reagents

pH Buffers

According to certain embodiments, the herein described formulations andcompositions for the reduction of sedimentation rate of one or moreerythrocytes in a blood sample include one or more pH buffers. In someembodiments, the pH buffer is any of a large number of compounds knownin the art for their ability to resist changes in the pH of a solution,such as in an aqueous solution in which the pH buffer is present.Selection of one or more particular pH buffers for inclusion in a stablestorage composition may be done based on the present disclosure andaccording to routine practices in the art, and may be influenced by avariety of factors including the pH that is desired to be maintained,the nature of the biological sample, the solvent conditions to beemployed, the other components of the formulation to be used, and othercriteria. For example, typically a pH buffer is employed at a pH that iswithin about 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0 pH unitof a proton dissociation constant (pKa) that is a characteristic of thebuffer.

Non-limiting examples of pH buffers include citric acid, tartaric acid,malic acid, sulfosalicylic acid, sulfoisophthalic acid, oxalic acid,borate, CAPS (3-(cyclohexylamino-1-propanesulfonic acid), CAPSO(3-(cyclohexylamino)-2-hydroxy-1-propanesulfonic acid), EPPS(4-(2-hydroxyethyl)-1-piperazinepropanesulfonic acid), HEPES(4-(2-hydroxyethyDpiperazine-1-ethanesulfonic acid), MES(2-(N-morpholino)ethanesulfonic acid), MOPS(3-(N-morpholino)propanesulfonic acid), MOPSO(3-morpholino-2-hydroxypropanesulfonic acid), PIPES(1,4-piperazinediethanesulfonic acid), TAPS(N-[tris(hydroxymethy)methyl]-3-aminopropanesulfonic acid), TAPSO(2-hydroxy-3-[tris(hydroxymethyl)methylamino]-1-propanesulfonic acid),TES (N-[tris(hydroxymethyl)methyl]-2-aminoethanesulfonic acid), bicine(N,N-bis(2-hydroxyethyl)glycine), tricine(N-[tris(hydroxymethyl)methyl]glycine), tris(tris(hydroxymethyl)aminomethane) and bis-tris(2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)-1,3-propanediol). Insome embodiments, the formulations have a pH of about 4.0, 4.1, 4.2,4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6,5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0,7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4,8.5, 8.6, 8.7, 8.8, 8.9, or 9.0.

Disaccharide Derivatives

In certain embodiments, the formulations or compositions for reductionof sedimentation rate of erythrocytes in a blood sample include at leastone halogenated disaccharide derivative. In some embodiments, thehalogenated disaccharide derivative is a di- or tri-chlorinateddisaccharide. In some embodiments, such di- or tri-chlorinateddisaccharides unexpectedly are capable of reducing sedimentation rate oferythrocytes in a blood sample either alone or in the presence of only abuffer. Halogenated disaccharide derivatives are known, e.g., see USPatent Publication No. 2014/0065062, and include sucralose(1,6-dichloro-1,6-dideoxy-β-D-fructofuranosyl-4-chloro-4-deoxy-α-D-galactopyranoside),trichloronated maltose,1,6-dichloro-1,6-dideoxy-β-D-fructofuranosyl-4-chloro-4-deoxy-6-O-monododecanoate-α-D-galactopyranoside,and1,6-dichloro-1,6-dodeoxy-β-D-fructofuranosyl-4-chloro-4-deoxy-6-O-monotetradecanoate-α-D-galactopyranoside.Selection of one or more particular halogenated disaccharide derivativefor inclusion in a formulation or composition for reduction ofsedimentation rate of erythrocytes in a blood sample may be done basedon the present disclosure and according to routine practices in the art,and may be influenced by a variety of factors including otherformulation components.

In some embodiments, the halogenated disaccharide derivative issucralose. In some embodiments, the sucralose is provided in solution asa formulation for mixing with a blood sample. In some embodiments, thesolution is an aqueous solution. In some embodiments, the sucralose ispresent in the formulation at about 5-500 mM. In some embodiments, thesucralose is present in the formulation at about 10-500 mM. In someembodiments, the sucralose is present in the formulation at about 50-500mM. In some embodiments, the sucralose is present in the formulation atabout 100-500 mM. In some embodiments, the sucralose is present in theformulation at about 250-500 mM. In some embodiments, the sucralose ispresent in the formulation at about 5-630 mM. In some embodiments, thesucralose is present in the formulation at about 5-750 mM. In someembodiments, the sucralose is present in the formulation at about 10-750mM. In some embodiments, the sucralose is present in the formulation atabout 50-750 mM. In some embodiments, the sucralose is present in theformulation at about 100-750 mM. In some embodiments, the sucralose ispresent in the formulation at about 250-750 mM. In some embodiments, theformulation is a mixture of water and sucralose.

In some embodiments, the formulation is provided in an amount sufficientto produce a final concentration of sucralose of about 5 to about 25 mM,when mixed with a blood sample. In some embodiments, the sucralose ispresent in the formulation at about 500 mM and is mixed with a bloodsample at a ratio of 1:20 (v/v) (formulation to blood). In someembodiments, the sucralose is present in the formulation at greater than25 mM up to 100 mM. In some embodiments, the sucralose is present in theformulation at about 13-24 mM. In some embodiments, the sucralose isprovided in powder form as a formulation for mixing with a blood sample.In some embodiments, the sucralose powder is provided in an amountsufficient to produce a final concentration of sucralose of about 5 toabout 25 mM, when mixed with a blood sample.

In some embodiments, the sucralose is present at a final concentrationof about 5-100 mM, when mixed with a blood sample. In some embodiments,the sucralose is present at a final concentration of about 5-50 mM, whenmixed with a blood sample. In some embodiments, the sucralose is presentat a final concentration of about 5-25 mM, when mixed with a bloodsample. In some embodiments, the sucralose is present at a finalconcentration of about 5 up to but not including 25 mM, when mixed witha blood sample. In some embodiments, the sucralose is present at a finalconcentration of about 5-20 mM, when mixed with a blood sample. In someembodiments, the sucralose is present at a final concentration of about5-15 mM, when mixed with a blood sample. In some embodiments, thesucralose is present at a final concentration of about 10-20 mM, whenmixed with a blood sample. In some embodiments, the sucralose is presentat a final concentration of about 10-15 mM, when mixed with a bloodsample. In some embodiments, the sucralose is present at a finalconcentration of about 5-10 mM, when mixed with a blood sample. In someembodiments, the sucralose and is present at a final concentration ofabout 25 mM, when mixed with a blood sample.

Anticoagulants

In some embodiments, an anticoagulant is included in the presentlydescribed formulations and compositions. Such anticoagulants are knownin the art. Exemplary anticoagulants include acid citrate dextrosesolution (ACD), ethylenediaminetetraacetic acid (EDTA), tri-potassiumethylenediaminetetraacetic acid (K₃EDTA), di-potassiumethylenediaminetetraacetic acid (K₂EDTA), heparin, sodium heparin,sodium fluoride, lithium heparin, sodium citrate, hirudin, and sodiumpolyanethol sulfonate (SPS). In some embodiments, the anticoagulant iscontained within a blood collection tube.

Exemplary Formulations for Reduction of Erythroctyte Sedimentation Ratein a Blood Sample

Described herein, in some embodiments, are formulations comprisingsucralose. In some embodiments, the sucralose is present at a finalconcentration of about 5 mM sucralose to about 50 mM sucralose, whenmixed with a blood sample. In some embodiments, the sucralose is presentat a final concentration of about 5 mM sucralose to about 25 mMsucralose, when mixed with a blood sample. In some embodiments, thesucralose is present at a final concentration of about 5 mM sucralose upto, but not including, 25 mM sucralose. In some embodiments, thesucralose is present at a final concentration of about 5 mM sucralose toabout 20 mM sucralose. In some embodiments, the sucralose is present ata final concentration of about 5 mM sucralose to about 15 mM sucralose.In some embodiments, the sucralose is present at a final concentrationof about 5 mM sucralose to about 10 mM sucralose. In some embodiments,the sucralose is present as a solution. In some embodiments, thesolution is an aqueous solution. In some embodiments, the sucralose ispresent as a powder. In some embodiments, the formulations furthercomprise an anticoagulant. In some embodiments, the anticoagulant isselected from the group consisting of acid citrate dextrose solution(ACD), sodium heparin, sodium fluoride, lithium heparin, tri-potassiumethylenediaminetetraacetic acid (K₃EDTA), di-potassiumethylenediaminetetraacetic acid (K₂EDTA), hirudin, and sodiumpolyanethol sulfonate (SPS). In some embodiments, the anticoagulant isacid citrate dextrose solution (ACD). In some embodiments, theanticoagulant is sodium heparin. In some embodiments, the anticoagulantis sodium fluoride. In some embodiments, the anticoagulant is lithiumheparin. In some embodiments, the anticoagulant is tri-potassiumethylenediaminetetraacetic acid (K₃EDTA). In some embodiments, theanticoagulant is di-potassium ethylenediaminetetraacetic acid (K₂EDTA).In some embodiments, the anticoagulant is hirudin. In some embodiments,the anticoagulant is sodium polyanethol sulfonate (SPS). In someembodiments, the formulation is contained within a blood collectiontube.

Methods for Preparing Formulations for Reducing ErythrocyteSedimentation Rate in a Blood Sample

Methods for preparing the formulations described herein for reduction oferythrocyte sedimentation rate in a blood sample employ techniques thatare well-known to those skilled in the art and generally usecommercially available reagents. In some embodiments, the formulationsare prepared as concentrated stock solutions of the formulationreagents, e.g., 2×, 5×, 10×, 20× or the like, so as to be admixed withthe blood sample at the appropriate ratios to produce the desired finalconcentrations of sucralose in the blood sample.

Compositions of Erythrocytes in a Blood Sample with ReducedSedimentation Rate

Described herein, in some embodiments, are compositions comprising ablood sample and sucralose, wherein the sucralose is at a concentrationof about 5 mM sucralose up to, but not including, about 25 mM sucralose.In some embodiments, the compositions further comprise an anticoagulant.In some embodiments, the anticoagulant is selected from the groupconsisting of acid citrate dextrose solution (ACD), sodium heparin,sodium fluoride, lithium heparin, tri-potassiumethylenediaminetetraacetic acid (K₃EDTA), di-potassiumethylenediaminetetraacetic acid (K₂EDTA), hirudin, and sodiumpolyanethol sulfonate (SPS). In some embodiments, the anticoagulant isacid citrate dextrose solution (ACD). In some embodiments, theanticoagulant is acid citrate dextrose solution A (ACD-A). In someembodiments, the anticoagulant is acid citrate dextrose solution B(ACD-B). In some embodiments, the anticoagulant is sodium heparin. Insome embodiments, the anticoagulant is sodium fluoride. In someembodiments, the anticoagulant is lithium heparin. In some embodiments,the anticoagulant is tri-potassium ethylenediaminetetraacetic acid(K₃EDTA). In some embodiments, the anticoagulant is di-potassiumethylenediaminetetraacetic acid (K₂EDTA). In some embodiments, theanticoagulant is hirudin. In some embodiments, the anticoagulant issodium polyanethol sulfonate (SPS). In some embodiments, the compositionis contained within a blood collection tube. In some embodiments, theblood collection tube is an evacuated blood collection tube. In someembodiments, the blood is collected from a subject. In some embodiments,the subject is a mammal. In some embodiments, the subject is a human. Insome embodiments, the compositions of one or more erythrocytes in ablood sample with reduced sedimentation rate are stored in theformulations described herein for extended periods of time beforeanalysis in, for example, a microfluidic device.

Articles of Manufacture

In certain embodiments, articles of manufacture are provided, whichcomprise a formulation provided herein, contained within a suitableblood collection tube, container or vessel for collection of abiological sample. In some embodiments, these articles of manufactureare used for reducing sedimentation rate of one or more erythrocytes ina blood sample at the time of biological sample collection. In certainembodiments, the blood collection tube is an evacuated blood tube havingless than atmospheric pressure to withdraw a predetermined volume ofwhole blood. In some embodiments, the blood collection tube containsabout 28.6 mg of sucralose powder and the blood collection tube is of asize to contain a blood draw volume of 3.0 mL blood to produce a finalsucralose concentration of about 24 mM after the addition of 3.0 mLblood. In some embodiments, the blood collection tube contains about33.4 mg of sucralose powder and the blood collection tube is of a sizeto contain a blood draw volume of 3.5 mL blood. In some embodiments, theblood collection tube contains about 42.9 mg of sucralose powder and theblood collection tube is of a size to contain a blood draw volume of 4.5mL blood. In some embodiments, the blood collection tube contains about52.4 mg of sucralose powder and the blood collection tube is of a sizeto contain a blood draw volume of 5.5 mL blood. In some embodiments, theblood collection tube contains about 95.4 mg of sucralose powder and theblood collection tube is of a size to contain a blood draw volume of 10mL blood. In some embodiments, these articles of manufacture are used inthe kits and methods described herein.

Kits

In certain embodiments, there are provided kits comprising any one ofthe articles of manufacture described herein and a package insert. Insome embodiments, the components of the kit are supplied in a container.In some embodiments, the container is a compartmentalized plasticenclosure. In some embodiments, the container includes a hermeticallysealable cover so that the contents of the kit can be sterilized andsealed for storage.

Methods for Reducing Sedimentation Rate of Erythrocytes in a BloodSample

Described herein, in some embodiments, are in vitro methods for reducingthe erythrocyte sedimentation rate in a blood sample, comprising:combining a sample of blood with an amount of a formulation comprisingsucralose, wherein the amount is sufficient to produce a treated bloodsample having a sucralose concentration of at least about 5 mMsucralose, thereby reducing the erythrocyte sedimentation rate ascompared to erythrocyte sedimentation rate in an untreated blood sample.In some embodiments, the treated blood sample has a sucraloseconcentration of about 5 mM sucralose to about 50 mM sucralose. In someembodiments, the treated blood sample has a sucralose concentration ofabout 5 mM sucralose to about 25 mM sucralose. In some embodiments, thetreated blood sample has a sucralose concentration of about 5 mMsucralose up to but not including 25 mM sucralose. In some embodiments,erythrocyte sedimentation rate is reduced by at least 10%, at least 20%,at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, atleast 80%, at least 90%, at least 95% of the erythrocyte sedimentationrate of the untreated blood sample. In some embodiments, the formulationis a powder. In some embodiments, the formulation is a solution. In someembodiments, the solution is an aqueous solution. In some embodiments,the formulation consists of sucralose. In some embodiments, theformulation further comprises an anticoagulant. In some embodiments, theanticoagulant is selected from the group consisting of acid citratedextrose solution (ACD), sodium heparin, sodium fluoride, lithiumheparin, tri-potassium ethylenediaminetetraacetic acid (K₃EDTA),di-potassium ethylenediaminetetraacetic acid (K₂EDTA), hirudin, andsodium polyanethol sulfonate (SPS). In some embodiments, theanticoagulant is acid citrate dextrose solution (ACD). In someembodiments, the anticoagulant is sodium heparin. In some embodiments,the anticoagulant is sodium fluoride. In some embodiments, theanticoagulant is lithium heparin. In some embodiments, the anticoagulantis tri-potassium ethylenediaminetetraacetic acid (K₃EDTA). In someembodiments, the anticoagulant is di-potassiumethylenediaminetetraacetic acid (K₂EDTA). In some embodiments, theanticoagulant is hirudin. In some embodiments, the anticoagulant issodium polyanethol sulfonate (SPS). In some embodiments, the formulationis contained within a blood collection tube, and the combining stepoccurs within the blood collection tube. In some embodiments, the bloodcollection tube is an evacuated blood collection tube. In someembodiments, the blood is collected from a subject. In some embodiments,the subject is a mammal. In some embodiments, the subject is a human.

Described herein, in some embodiments, are methods for maintaining oneor more erythrocytes in suspension in a blood sample, comprising:combining a sample of blood with an amount of a formulation comprisingsucralose, wherein the amount is sufficient to produce a treated bloodsample having a sucralose concentration of at least about 5 mMsucralose, thereby maintaining the one or more erythrocytes insuspension for a period of at least 30 minutes as compared to anuntreated blood sample. In some embodiments, the one or moreerythrocytes remain in suspension for a period of at least 10 minutes,at least 20 minutes, at least 30 minutes, at least 40 minutes, at least50 minutes, at least 1 hour, at least 2 hours, at least 3 hours, atleast 4 hours, at least 5 hours, at least 6 hours, at least 7 hours, atleast 8 hours, at least 10 hours, at least 12 hours, at least 24 hoursor at least 48 hours. In some embodiments, the treated blood sample hasa sucralose concentration of about 5 mM sucralose to about 25 mMsucralose. In some embodiments, the treated blood sample has a sucraloseconcentration of about 5 mM sucralose up to but not including 25 mMsucralose. In some embodiments, at least 10%, at least 20%, at least30%, at least 40%, at least 50%, at least 60%, at least 70%, at least80%, at least 90%, at least 95% of the one or more erythrocytes remainin suspension in the treated blood sample as compared to the untreatedblood sample. In some embodiments, the formulation is a powder. In someembodiments, the formulation is a solution. In some embodiments, thesolution is an aqueous solution. In some embodiments, the formulationconsists of sucralose. In some embodiments, the formulation furthercomprises an anticoagulant. In some embodiments, the anticoagulant isselected from the group consisting of acid citrate dextrose solution(ACD), sodium heparin, sodium fluoride, lithium heparin, tri-potassiumethylenediaminetetraacetic acid (K₃EDTA), di-potassiumethylenediaminetetraacetic acid (K₂EDTA), hirudin, and sodiumpolyanethol sulfonate (SPS). In some embodiments, the anticoagulant isacid citrate dextrose solution (ACD). In some embodiments, theanticoagulant is sodium heparin. In some embodiments, the anticoagulantis sodium fluoride. In some embodiments, the anticoagulant is lithiumheparin. In some embodiments, the anticoagulant is tri-potassiumethylenediaminetetraacetic acid (K₃EDTA). In some embodiments, theanticoagulant is di-potassium ethylenediaminetetraacetic acid (K₂EDTA).In some embodiments, the anticoagulant is hirudin. In some embodiments,the anticoagulant is sodium polyanethol sulfonate (SPS). In someembodiments, the formulation is contained within a blood collectiontube, and the combining step occurs within the blood collection tube. Insome embodiments, the blood collection tube is an evacuated bloodcollection tube. In some embodiments, the blood sample is collected froma subject. In some embodiments, the subject is a mammal. In someembodiments, the subject is a human.

Blood collection tubes, bags, containers and vessels are well-known inthe art and have been employed by medical practitioners for decades.Blood collected for reduction of erythrocyte sedimentation rate may beobtained using any method or apparatus commonly employed by thoseskilled in the art such as venipuncture or finger prick. In someembodiments, when the blood is collected by venipuncture, theformulation is located inside the blood collection tube, e.g., anevacuated tube (VACUTAINER® blood collection tube, Becton Dickinson orVACUETTE® blood collection tube, Greiner Bio-One) at the time that theblood sample is obtained from the subject. In some embodiments, when theblood is collected by venipuncture, the formulations are added to analready obtained whole blood sample, either immediately or shortly afterit is withdrawn.

In some embodiments, the methods as described herein use the articles ofmanufacture and kits disclosed.

The following Examples are presented by way of illustration and notlimitation.

Example 1 Reduction of Erythrocyte Sedimentation Rate in Whole Blood byAddition of Sucralose

This Example describes reduction of erythrocyte sedimentation in wholeblood by addition of sucralose.

Fresh blood was collected into BD K₂EDTA VACUTAINER® tubes and pooled.Blood and a solution of 0.5 M sucralose were mixed at a ratio of 20:1 byaliquotting 952 μL of fresh blood into 2 mL centrifuge tubes containing48 μL of formulation, resulting in a concentration of about 24 mMsucralose. 48 μL of PBS and 0.9% saline served as controls. Blood withno formulation (NF) added served as an additional control. The filledcentrifuge tubes were gently inverted five times to mix and storedupright on the benchtop at approximately 25° C. Tubes were photographedat 0 hours (FIG. 1A), 6 hours (FIG. 1B), and 24 hours (FIG. 1C) againsta white background for visual analysis of sedimentation rate.

As shown in FIGS. 1A-C, whole blood collected in K₂EDTA tubes containingsucralose had lower sedimentation rate of the erythrocytes after storageof the aliquots at ambient temperature for 6 hours and 24 hours comparedto whole blood only (NF) or whole blood following addition of either PBSor saline.

Example 2 Effect of Sucralose Concentration and Other Saccharides onReduction of Erythrocyte Sedimentation Rate

This Example illustrates the effect on sucralose concentration and othersaccharides on reduction of erythrocyte sedimentation rate in wholeblood.

Fresh blood was collected into BD K₂EDTA VACUTAINER® tubes and pooled.952 ΞL of fresh blood was aliquot into 2 mL centrifuge tubes, eachcontaining 48 μL of sucralose at the indicated concentration in FIG. 2,or the indicated saccharide (ML848, a di-chlorinated monosaccharide, orDG783, a mono-fluorinated monosaccharide) at the indicatedconcentration. Sucralose solutions of different concentrations wereprepared by dilution of 0.5 M sucralose to 10 mM in water. Formulationswere adjusted to 300 mOsmol with NaCl, with the exception offormulations of the highest concentration that had an osmolarity of 500mOsmol. 48 μL of PBS and 0.9% saline served as controls. Finalconcentrations of sucralose or the indicated saccharide shown in FIG. 2were as follows (from left to right): 24 mM sucralose, 12 mM sucralose,6.24 mM sucralose, 2.88 mM sucralose, 1.44 mM sucralose, 0.96 mMsucralose, 0.48 mM sucralose, 24 mM sucrose, 2.4 mM sucrose, 24 mMtrehalose, 24 mM melezitose, 4.8 mM ML848, and 24 mM DG783.

The filled centrifuge tubes were gently inverted five times to mix andstored upright on the benchtop at approximately 25° C. Tubes werephotographed at 1 hour (FIG. 2, upper panel) and 6 hours (FIG. 2, centerpanel) against a white background for visual analysis of sedimentationrate. After 7 hours of storage, tubes were centrifuged for 20 min at3000 rpm to determine the effect of hemolysis on sedimentation rate byvisual analysis (FIG. 2, lower panel).

FIG. 2 shows that whole blood collected in K₂EDTA tubes containing highconcentrations of sucralose had lower sedimentation rate of erythrocytesafter storage of aliquots at ambient temperature for 1 hour (upperpanel) and 6 hours (center panel). Lower sedimentation rate oferythrocytes was not observed in samples that had final sucraloseconcentrations below 5 mM, or in samples following addition of sucrose,trehalose, melezitose, ML848, DG783, PBS, or saline to whole blood.

Homogeneity of whole blood in the presence of a high concentration ofsucralose was not due to excessive hemolysis of erythrocytes, as shownby centrifugation of the sample after 7 hours of storage (FIG. 2, lowerpanel). Centrifugation resulted in clear separation of plasma withoutcoloration of the plasma layer similar to that seen for the saline andPBS sample controls (FIG. 2, lower panel). Significant hemolysis was notseen at final sucralose concentrations at or below 40 mM.

Example 3 Effect of Sucralose Added as a Powder on Reduction ofErythrocyte Sedimentation Rate

This Example describes the effect of addition of sucralose in powderform on reduction of erythrocyte sedimentation rate in whole blood.

Fresh blood was collected into BD K₂EDTA VACUTAINER® tubes and pooled. 1mL of fresh blood was aliquotted into 2 mL centrifuge tubes containingsucralose powder. Final concentrations of sucralose ranged from 7.5 mMto 100 mM, corresponding to 3.0 mg to 39.8 mg per mL of blood. The noformulation (NF) control sample received no addition of sucralose. Tubeswere inverted at least five times until no visible undissolved materialremained at the bottom of the tubes. Tubes were stored upright on thebenchtop at approximately 25° C. Tubes were photographed at 1 hour (FIG.3, upper panel) and 6 hours (FIG. 3, lower panel) against a whitebackground for visual analysis of sedimentation rate.

Data in FIG. 3 shows that the rate of erythrocyte sedimentation wasinversely proportional to the concentration of sucralose. At a sucraloseconcentration of 24 mM or greater, homogeneity of whole blood samplesstored at ambient temperatures for 6 hours was observed, while lowerconcentrations of sucralose resulted in separation of the erythrocyteand plasma layers.

Example 4 Effect of Blood Collection Conditions on Reduction ofErythrocyte Sedimentation Rate

This Example describes the effect of different anticoagulants present atthe time of blood collection on reduction of erythrocyte sedimentationrate in whole blood.

To screen for collection conditions, fresh blood was collected into aseries of BD VACUTAINER® or Greiner Bio-One VACUETTE® low-volume bloodcollection tubes containing different anticoagulants, including acidcitrate dextrose solution B (ACD-B), tri-potassiumethylenediaminetetraacetic acid (K₃EDTA), sodium heparin (NaHep),lithium heparin (LiHep), sodium fluoride (NaF), and sodium polyanetholsulfonate (SPS). Collection into tubes containing di-potassiumethylenediaminetetraacetic acid (K₂EDTA) served as a control. 952 μL offresh blood was aliquot into 2 mL centrifuge tubes containing 48 μL of0.5 M sucralose or PBS. No formulation (NF) added served as anadditional control. The filled centrifuge tubes were gently invertedfive times to mix and stored upright on the benchtop at approximately25° C. Tubes were photographed at 2 hours (FIG. 4, upper panel) and 8hours (FIG. 4, lower panel) against a white background for visualanalysis of sedimentation rate. As shown in FIG. 4, addition of 0.5 Msucralose, for a final sucralose concentration of 24 mM in the treatedsample, resulted in reduction of erythrocyte sedimentation rate for eachanticoagulant present in the blood collection tube at the time of bloodcollection. By contrast, addition of PBS had no effect on erythrocytesedimentation rate.

Example 5 Effect of Sucralose Compared to Polyols and HalogenatedPolyols on Reduction of Erythrocyte Sedimentation Rate

This Example describes the effect of sucralose compared to the effect ofpolyols and halogenated polyols on reduction of erythrocytesedimentation rate in whole blood.

Fresh blood was collected into BD K₂EDTA VACUTAINER® tubes and pooled.952 μL of fresh blood was aliquot into 2 mL centrifuge tubes containing48 μL of a solution of 0.5 M sucralose, the indicated polyol, or theindicated halogenated polyol. Additives at 0.25 M were adjusted to 300mOsmol with NaCl. PBS and no formulation (NF) added served as controls.The filled centrifuge tubes were gently inverted five times to mix andstored upright on the benchtop at approximately 25° C. Tubes werephotographed at 2 hours against a white background for visual analysisof sedimentation rate.

Data in FIG. 5 shows that addition of a solution of 0.5M sucralose, fora final sucralose concentration of 24 mM in the treated sample, resultedin reduced erythrocyte sedimentation rate. By contrast, addition ofindicated polyols or halogenated polyols had no apparent effect onerythrocyte sedimentation.

Unless the context requires otherwise, throughout the presentspecification and claims, the word “comprise” and variations thereof,such as, “comprises” and “comprising,” which is used interchangeablywith “including,” “containing,” or “characterized by,” is inclusive oropen-ended language and does not exclude additional, unrecited elementsor method steps. The phrase “consisting of” excludes any element, step,or ingredient not specified in the claim. The phrase “consistingessentially of” limits the scope of a claim to the specified materialsor steps and those that do not materially affect the basic and novelcharacteristics of the claimed invention. The present disclosurecontemplates embodiments of the invention compositions and methodscorresponding to the scope of each of these phrases. Thus, a compositionor method comprising recited elements or steps contemplates particularembodiments in which the composition or method consists essentially ofor consists of those elements or steps.

Reference throughout this specification to “one embodiment” or “anembodiment” or “an aspect” means that a particular feature, structure orcharacteristic described in connection with the embodiment is includedin at least one embodiment of the present invention. Thus, theappearances of the phrases “in one embodiment” or “in an embodiment” invarious places throughout this specification are not necessarily allreferring to the same embodiment. Furthermore, the particular features,structures, or characteristics may be combined in any suitable manner inone or more embodiments.

The various embodiments described above can be combined to providefurther embodiments. These and other changes can be made to theembodiments in light of the above-detailed description. In general, inthe following claims, the terms used should not be construed to limitthe claims to the specific embodiments disclosed in the specificationand the claims, but should be construed to include all possibleembodiments along with the full scope of equivalents to which suchclaims are entitled. Accordingly, the claims are not limited by thedisclosure.

While preferred embodiments of the present invention have been shown anddescribed herein, it will be obvious to those skilled in the art thatsuch embodiments are provided by way of example only. Numerousvariations, changes, and substitutions will now occur to those skilledin the art without departing from the invention. It should be understoodthat various alternatives to the embodiments of the invention describedherein may be employed in practicing the invention. It is intended thatthe following claims define the scope of the invention and that methodsand structures within the scope of these claims and their equivalents becovered thereby.

What is claimed is:
 1. A blood collection tube comprising a formulationfor reducing erythrocyte sedimentation rate, the formulation comprisingan anticoagulant and sucralose.
 2. The blood collection tube of claim 1,wherein the sucralose is present in an amount sufficient to provide afinal concentration of about 5.0 mM to about 100 mM sucralose uponaddition of the blood sample.
 3. The blood collection tube of claim 1,wherein the sucralose is present in an amount sufficient to provide afinal concentration of about 5.0 mM to about 50 mM sucralose uponaddition of the blood sample.
 4. The blood collection tube of claim 1,wherein the sucralose is present in an amount sufficient to provide afinal concentration of about 5.0 to about 25 mM sucralose upon additionof the blood sample.
 5. The blood collection tube of claim 1, whereinthe anticoagulant is selected from the group consisting of acid citratedextrose solution (ACD), sodium heparin, sodium fluoride, lithiumheparin, tri-potassium ethylenediaminetetraacetic acid (K₃EDTA),di-potassium ethylenediaminetetraacetic acid (K₂EDTA), hirudin, andsodium polyanethol sulfonate (SPS).
 6. The blood collection tube ofclaim 1, wherein the formulation is in the form of a powder, a solid, alyophilized form, or a solution.
 7. The blood collection tube of claim1, wherein the tube is an evacuated blood tube.
 8. The blood collectiontube of claim 1, wherein the formulation consists essentially of ananticoagulant and sucralose.
 9. The blood collection tube of claim 1,wherein the formulation reduces the erythrocyte sedimentation rate by atleast 10% compared to the erythrocyte sedimentation rate of an untreatedblood sample.
 10. The blood collection tube of claim 1, wherein theformulation reduces the erythrocyte sedimentation rate by at least 25%compared to the erythrocyte sedimentation rate of an untreated bloodsample.
 11. The blood collection tube of claim 1, wherein theformulation reduces the erythrocyte sedimentation rate by at least 50%compared to the erythrocyte sedimentation rate of an untreated bloodsample.
 12. The blood collection tube of claim 1, wherein theformulation reduces the erythrocyte sedimentation rate during storage ofthe sample over at least 24 hours compared to the erythrocytesedimentation rate of an untreated blood sample.
 13. The bloodcollection tube of claim 1, wherein the formulation reduces theerythrocyte sedimentation rate during storage of the sample over atleast 12 hours compared to the erythrocyte sedimentation rate of anuntreated blood sample.
 14. The blood collection tube of claim 1,wherein the formulation reduces the erythrocyte sedimentation rateduring storage of the sample over at least 6 hours compared to theerythrocyte sedimentation rate of an untreated blood sample.
 15. Theblood collection tube of claim 1, wherein the formulation reduces theerythrocyte sedimentation rate during storage of the sample at ambienttemperature compared to the erythrocyte sedimentation rate of anuntreated blood sample.
 16. The blood collection tube of claim 1,wherein at least about 2.0 mg of the sucralose is present in the bloodcollection tube per ml of added blood sample.